Two fates of monocyte differentiation are easily identified and separable: many monocytes develop into macrophages and others become dendritic cells (DCs). One of the differences between these populations is distinct migratory behavior; macrophages are much more sessile than DCs. We aim in the long term to understand what determines whether monocytes differentiate into one of these fates instead of the other in a given tissue, in order to learn to manipulate monocyte fate for regulation of immune responses and chronic inflammatory conditions. Our recent demonstration that the conversion of monocytes to sessile vs. migratory fates affects the progression of atherosclerosis highlights the importance of this pursuit. Moreover, it is still not clear how significantly monocytes account for DCs that emigrate to lymph nodes under homeostatic or inflammatory conditions. A stronger fundamental understanding of normal monocyte differentiation and trafficking is needed. We will trace monocyte differentiation to DCs vs. macrophages in the steady state using a cre recombinase-expressing mouse strain to follow monocytes in homeostasis, and we will directly analyze "pseudoafferent" lymph to trace monocyte-derived cells that enter peripheral lymphatic vessels (aim 1). We have also recently developed a technique to introduce a phagocytic label into circulating monocytes. This technique permits us to interogate monocyte fate in situ, including the mechanisms and consequences of conversion from one monocyte subset to another (aim 2). The use of this tracing technique also surprisingly revealed that some monocytes freshly exiting the bone marrow can have previously either derived from or have engulfed other leukocytes that recently entered the bone marrow from the periphery. Thus, monocytes which appear unactivated/"naive" in the blood can be antigen-experienced before leaving the bone marrow. More work is needed to unravel this surprising finding and understand its impact on the immune system (aim 3).